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Molecular control of kinetochore-microtubule dynamics and chromosome oscillations

Nature Cell Biology volume 12, pages 319–329 (2010)Cite this article

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Abstract

Chromosome segregation in metazoans requires the alignment of sister kinetochores on the metaphase plate. During chromosome alignment, bioriented kinetochores move chromosomes by regulating the plus-end dynamics of the attached microtubules. The bundles of kinetochore-bound microtubules alternate between growth and shrinkage, leading to regular oscillations along the spindle axis. However, the molecular mechanisms that coordinate microtubule plus-end dynamics remain unknown. Here we show that centromere protein (CENP)-H, a subunit of the CENP-A nucleosome-associated and CENP-A distal complexes (CENP-A NAC/CAD), is essential for this coordination, because kinetochores lacking CENP-H establish bioriented attachments but fail to generate regular oscillations, as a result of an uncontrolled rate of microtubule plus-end turnover. These alterations lead to rapid erratic movements that disrupt metaphase plate organization. We also show that the abundance of the CENP-A NAC/CAD subunits CENP-H and CENP-I dynamically change on individual sister kinetochores in vivo, because they preferentially bind the sister kinetochore attached to growing microtubules, and that one other subunit, CENP-Q, binds microtubules in vitro. We therefore propose that CENP-A NAC/CAD is a direct regulator of kinetochore-microtubule dynamics, which physically links centromeric DNA to microtubule plus ends.

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Figure 1: Loss of CENP-H abolishes sister-kinetochore oscillations and disrupts metaphase plate alignment.
Figure 2: Loss of CENP-H abrogates MT flux and abolishes control of kMT turnover.
Figure 3: Loss of CENP-H increases kMT stability.
Figure 4: CENP-A NAC/CAD regulates kMT dynamics independently of Aurora B/KMN.
Figure 5: CENP-H and CENP-I bind differentially to kinetochores attached to growing kinetochore fibres.
Figure 6: CENP-I accumulates preferentially on the sister kinetochore bound to growing MTs.
Figure 7: The CENP-A NAC/CAD subunit CENP-Q makes direct physical interactions with MTs.
Figure 8: Model for the function of the CENP-A NAC/CAD in controlling kMT dynamics.

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Acknowledgements

We thank the ETH Light Microscopy Centre and Yves Barral for microscopy support; Anna Akhmanova, Helder Maiato, Michel Steinmetz, Daniel Gerlich, Erich Nigg, Stephan Diekmann and Jennifer Lippincot-Schwarz for reagents; Jason Swedlow, Gaudenz Danuser for the joint development of the kinetochore tracking assay; Satyarebala Borusu for the EGFP–CENP-O cell line; Jennifer Winter for initial photoactivation observations; Itsaso Olasagasti for helping with the CENP-I intensity calculations; Kunyoshi Kaseda for help in analysing the photoactivation experiments; and Yves Barral, Monica Gotta, Helder Maiato, Jonathon Pines and members of the Barral, McAinsh and Meraldi laboratories for helpful discussions. Work in the McAinsh laboratory was supported by Marie Curie Cancer Care (A.D.M. and C.P.S.) and by a Fundação para a Ciência e Tecnologia fellowship (C.P.S.). A.C.A. is a member of the Life Science Zurich Graduate School in Molecular Life Sciences. Work in the Meraldi laboratory (A.C.A, R.H. and P.M.) was supported by a SNSF-Förderungprofessur and a EURYI award.

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Authors and Affiliations

  1. Institute of Biochemistry, Eidgenössische Technische Hochschule (ETH) Zurich, CH-8093, Zurich, Switzerland

    Ana C. Amaro, René Holtackers & Patrick Meraldi

  2. Chromosome Segregation Laboratory, Marie Curie Research Institute, The Chart, Oxted, UK

    Catarina P. Samora & Andrew D. McAinsh

  3. Department of Biology, University of Pennsylvania, Philadelphia, 19104, Pennsylvania, USA

    Enxiu Wang & Michael Lampson

  4. London Research Institute, Lincoln's Inn Fields Laboratories, Cancer Research UK, London, UK

    Isabel J. Kingston

  5. Molecular Motors Group, Marie Curie Research Institute, The Chart, Oxted, RH8 0TL, Surrey, UK

    Maria Alonso

  6. Centre for Mechanochemical Cell Biology, Warwick Medical School, University of Warwick, CV4 7AL, Coventry, UK

    Catarina P. Samora, Maria Alonso & Andrew D. McAinsh

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  1. Ana C. Amaro
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Contributions

Project conception, planning and data analyses were performed by A.C.A., C.P.S., A.D.M. and P.M. A.C.A. performed all experiments except the following: C.P.S. performed the photoactivation experiments and the biochemical experiments. R.H. generated the photoactivatable GFP–α-tubulin/H2B–mRFP cell line. E.W. and M.L. measured Aurora B activity. I.K. and M.A. contributed to the biochemical experiments. The manuscript was prepared by A.D.M. and P.M. with contributions by A.C.A. and C.P.S.

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Correspondence to Andrew D. McAinsh or Patrick Meraldi.

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Amaro, A., Samora, C., Holtackers, R. et al. Molecular control of kinetochore-microtubule dynamics and chromosome oscillations. Nat Cell Biol 12, 319–329 (2010). https://doi.org/10.1038/ncb2033

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